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Antimicrobial Agents and Chemotherapy, July 2007, p. 2436-2444, Vol. 51, No. 7
0066-4804/07/$08.00+0     doi:10.1128/AAC.01115-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Steady-State Disposition of the Nonpeptidic Protease Inhibitor Tipranavir when Coadministered with Ritonavir

Linzhi Chen,^1* John P. Sabo,¹ Elsy Philip,¹ Yanping Mao,¹ Stephen H. Norris,¹ Thomas R. MacGregor,¹ Jan M. Wruck,³ Sandra Garfinkel,² Mark Castles,¹ Amy Brinkman,⁴ and Hernan Valdez²

Departments of Drug Metabolism and Pharmacokinetics,¹ Virology,² Biometrics and Data Management, Boehringer Ingelheim Pharmaceuticals, Inc., Ridgefield, Connecticut,³ Covance Laboratories Inc., Madison, Winsconsin⁴

Received 4 September 2006/ Returned for modification 28 January 2007/ Accepted 30 April 2007

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The pharmacokinetic and metabolite profiles of the antiretroviral agent tipranavir (TPV), administered with ritonavir (RTV), in nine healthy male volunteers were characterized. Subjects received 500-mg TPV capsules with 200-mg RTV capsules twice daily for 6 days. They then received a single oral dose of 551 mg of TPV containing 90 µCi of [¹⁴C]TPV with 200 mg of RTV on day 7, followed by twice-daily doses of unlabeled 500-mg TPV with 200 mg of RTV for up to 20 days. Blood, urine, and feces were collected for mass balance and metabolite profiling. Metabolite profiling and identification was performed using a flow scintillation analyzer in conjunction with liquid chromatography-tandem mass spectrometry. The median recovery of radioactivity was 87.1%, with 82.3% of the total recovered radioactivity excreted in the feces and less than 5% recovered from urine. Most radioactivity was excreted within 24 to 96 h after the dose of [¹⁴C]TPV. Radioactivity in blood was associated primarily with plasma rather than red blood cells. Unchanged TPV accounted for 98.4 to 99.7% of plasma radioactivity. Similarly, the most common form of radioactivity excreted in feces was unchanged TPV, accounting for a mean of 79.9% of fecal radioactivity. The most abundant metabolite in feces was a hydroxyl metabolite, H-1, which accounted for 4.9% of fecal radioactivity. TPV glucuronide metabolite H-3 was the most abundant of the drug-related components in urine, corresponding to 11% of urine radioactivity. In conclusion, after the coadministration of TPV and RTV, unchanged TPV represented the primary form of circulating and excreted TPV and the primary extraction route was via the feces.

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Tipranavir (TPV) is a nonpeptidic (15, 16) sulfonamide-substituted dihydropyrone (Fig. 1) protease inhibitor (PI) marketed for the treatment of human immunodeficiency virus (HIV)-infected, treatment-experienced patients under the trade name Aptivus. Most viruses resistant to PIs retain susceptibility to TPV (1, 6, 12). Like other PIs, TPV binds directly to HIV aspartyl protease, thereby disrupting the catalytic site of the enzyme and preventing the protease-dependent cleavage of HIV gag and gag-pol polyproteins into smaller functional proteins (10, 15, 16). Clinical studies have demonstrated the significant activity of TPV against HIV type 1 in infected patients receiving twice-daily doses ranging from 300 to 1,200 mg (2, 3, 4, 9).

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FIG. 1. Structure of the sulfonamide-substituted dihydropyrone PI TPV. The asterisk denotes the 14C label.

In clinical studies with healthy volunteers and HIV-infected patients, TPV pharmacokinetic parameters, including the steady-state trough concentration, the area under the concentration-time curve (AUC) for plasma, the maximum concentration in plasma during the steady state (C_max), and the apparent terminal half-life (t_1/2), were substantially improved with the concomitant administration of ritonavir (RTV) (8, 9). Since RTV strongly inhibits cytochrome P450 3A4 (CYP3A4) (5), the boosted levels of TPV in plasma with the coadministration of TPV and RTV (TPV/r) indicate that the metabolism of TPV occurs via the cytochrome P450 (CYP450) pathway (8, 9). Phase II studies have also demonstrated that TPV is an inducer of CYP450 metabolism or intestinal P glycoprotein efflux, thereby lowering systemic RTV concentrations when administered with RTV (8, 11, 14). Clinical studies with healthy volunteers and HIV-infected patients have evaluated potential drug interactions of TPV/r and established that no TPV dose adjustments are required when TPV/r is administered in conjunction with other commonly employed antiretroviral agents (13; F. D. Goebel, J. P. Sabo, T. R. MacGregor, D. L. Mayers, and S. McCallister, presented at the HIV DART Conference, 2002).

The purpose of the present study was to characterize the pharmacokinetic and metabolite profiles of TPV when administered with RTV.

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Materials. TPV, [¹⁴C]TPV, and TPV glucuronide were synthesized at Boehringer Ingelheim Pharmaceuticals, Inc. (Ridgefield, CT). The position of the ¹⁴C label is indicated in Fig. 1. The chemical identities of the compounds were established by high-performance liquid chromatography, mass spectrometry, and nuclear magnetic resonance. The radiochemical purity of [¹⁴C]TPV was determined by liquid chromatography (LC)-radiochromatography to be >98%. Aquasol-2, Ultima Gold XR, and Ultima FLO-M scintillation cocktails were purchased from PerkinElmer Life and Analytical Sciences (Shelton, CT). All other chemicals were provided by standard commercial sources and were of reagent grade or better.

Subjects, dosing, and sample collection. This phase I, open-label, multiple-oral-dose study was conducted to characterize the pharmacokinetics of TPV and its metabolites, to elucidate the disposition of the parent compound and the total radioactivity by excretion and mass balance during the steady state, and to identify and quantify major metabolites of TPV in plasma, urine, and feces. The study was conducted in accordance with the Nuclear Regulatory Commission regulations and the Declaration of Helsinki (1964 and subsequent revisions). Of the 12 healthy male volunteers enrolled, 9 completed all phases of the study, including the administration of [¹⁴C]TPV/r. Subjects were nonsmokers and received standardized meals and the same supervised doses of TPV/r, i.e., 500 mg of TPV plus 200 mg of RTV twice daily for 6 days. On day 7, participants received a light snack at 6:00 am, 60 min prior to the administration of 500 mg of [¹⁴C]TPV (90 µCi) plus 200 mg of RTV. Subjects fasted for an additional 4 h after the dose administration. At 7:00 pm that evening, participants resumed dosing with nonradioactive TPV at 500 mg and RTV at 200 mg twice daily and continued on this dosing regimen up to day 20 of the study. Subjects did not receive any medications known to affect CYP450 activity, and coffee, tea, cola, chocolate, St. John's wort, milk thistle, garlic supplements, Seville oranges, grapefruit, grapefruit juice, and alcohol were avoided during the course of the study.

Blood samples were collected daily, immediately prior to the evening doses of TPV/r on study days 1 through 7 (5 ml) and prior to the morning doses on study days 8 through 15 (10 ml). On day 7, serial 10-ml blood samples were obtained 10 min prior to the radioactive dose and at 0.5, 1, 1.5, 2, 3, 4, 5, 6, 8, 10, 12, and 24 h post-radioactive dose. Both whole blood and plasma were analyzed for radioactivity. Additional 10-ml blood samples for metabolite profiling were collected at 3, 8, and 12 h post-radioactive dose. The 3-h time point coincides with the mean peak plasma TPV concentration (8), while the 8- and 12-h time points were chosen for metabolite profiling as late time points near the end of the 12-h dosing interval. Plasma was prepared immediately after sampling by centrifugation at 5°C for 10 min at 2,000 x g.

After the administration of the radioactive dose, urine and feces were collected until the end of the study. All samples were stored at approximately –20°C until analyzed.

Radioactivity measurement. Feces and tissue samples were homogenized in water and a mixture of ethanol and water (1:1), respectively, which were approximately three to five times the sample weight. Whole-blood, fecal homogenate, and tissue homogenate samples were combusted before radioactivity counting with a Packard 307 sample oxidizer (Packard Instrument Co., Meriden, CT), and the resulting ¹⁴CO₂ was trapped in a mixture of Perma Fluor and Carbo-Sorb (Packard Instrument Co., Meriden, CT). Aliquots of samples (0.2 ml of blood and plasma, 0.5 ml of urine, and 0.5 g of fecal and tissue homogenates) were mixed with 5 ml of Ultima Gold XR scintillation cocktail and evaluated for radioactivity by using Packard 2900TR liquid scintillation counters (Packard Instrument Co.). Scintillation counting data (expressed in counts per minute) were automatically corrected for counting efficiency by using the external standardization technique and an instrument-stored quench curve generated from a series of sealed quenched standards.

Quantitation of TPV in plasma. The concentrations of TPV in plasma were determined using a validated LC-tandem mass spectrometry (MS-MS) method. Plasma samples were prepared by a two-step liquid-liquid extraction procedure; the use of an ethyl acetate-hexane mixture was followed by a hexane wash. Chromatographic separation was achieved using a 2.0- by 30-mm, 4-µm-particle-size Synergi Polar-RP column (Phenomenex Inc., Torrance, CA). The mobile phase consisted of 50% 20 mM formic acid-10 mM acetic acid and 50% acetonitrile; the run time was 6 min. The detector used was a MicroMass QuattroLC triple-quadrupole mass spectrometer (Waters, Milford, MA), which was equipped with an electrospray ion source and operated in positive-selective-reaction monitoring mode. Key MS-MS operating parameters included the following: capillary voltage, 1 kV; nebulizer gas, 110 liters/h; desolvation gas, 1,300 liters/h; desolvation temperature, 300°C; cone voltage, 30 V; and collision offset, 20 V. Of the two overlapping calibration curves used in this study, one had a calibration range of 25.0 to 2,000 ng/ml and the other had a calibration range of 1,000 to 20,000 ng/ml. Two calibration curves with different calibration ranges were used to minimize sample manipulation during bioanalysis. All of the samples were assayed with the higher-range curve first. For the samples that had a concentration of less than 1,000 ng/ml (the lower limit of quantitation of the higher-range curve), there was no reportable result, and these samples were subsequently reassayed with the lower-range curve.

Metabolite sample extraction. The plasma samples collected from the nine subjects at each of the three time points, 3, 8, and 12 h postdose, were pooled for metabolite profiling and identification. A 20- to 25-ml aliquot of the plasma pool was evaporated at room temperature under a nitrogen stream in a Zymark TurboVap LV evaporator (Zymark, Hopkinton, MA), and the residue was then extracted four to six times with 10 to 20 ml of methanol. For each extraction, the mixture was sonicated for 5 min, and the supernatant was separated by centrifugation. The extracts were combined, evaporated with the TurboVap, and subjected to further cleanup via multiple evaporation-extraction cycles with methanol and acetonitrile. The final sample was reconstituted in 0.30 to 0.43 ml of methanol and transferred to an autosampler vial for LC-radiochromatography-MS-MS analysis. The total extraction recovery of radioactivity from plasma ranged from 87.8 to 100.8%.

The urine samples collected from each subject during the first 24 h after [¹⁴C]TPV dosing were pooled. The majority (50.0 to 63.3%) of urine radioactivity was found in the urine from the 0-to-24-h sample collection period, and the remaining radioactivity was spread throughout large volumes of urine across several days. Equal percentages (by volume) of the 0-to-24-h urine samples were combined to make the pools used for metabolite profiling and identification. To a 100-ml aliquot of pooled urine was added 6 ml of 28 to 30% ammonium hydroxide. The urine samples were then extracted via solid-phase extraction with 35-cm³/6-g Oasis HLB solid-phase extraction cartridges (Waters Corporation, Millford, MA). After loading, the solid-phase extraction cartridge was washed with 25 to 35 ml of 2% ammonium hydroxide, and the sample was eluted with methanol. The eluate was evaporated and further cleaned up via multiple evaporation-extraction cycles with acetonitrile. The final samples were reconstituted in 0.4 to 0.5 ml of methanol and transferred to an autosampler vial for metabolite profiling and identification. The total extraction recovery of radioactivity from urine ranged from 84.1 to 101.7%.

A fecal pool for each subject for metabolite profiling and identification was also prepared by combining aliquots of fecal homogenates from three to five sampling time points at equal percentages (by volume). Depending on the subject, feces from 0 to 24 h up to 120 to 144 h were pooled. The pools corresponded to 90% or more of the total radioactivity ultimately excreted in feces. An 18- to 30-ml aliquot of the pooled fecal homogenate was centrifuged, the supernatant was discarded, and the pellet was extracted with acetonitrile, methanol, and acidified methanol. The extracts were combined, evaporated, and further cleaned up via multiple evaporation-extraction cycles with acetonitrile. The final samples were reconstituted in 0.3 to 0.5 ml of methanol and then transferred to an autosampler vial for metabolite profiling and identification. The total extraction recovery of radioactivity from feces ranged from 81.2 to 106.1%.

Metabolite profiling and identification. Metabolite profiling and identification was carried out with an LC-radiochromatography-MS-MS system, which consisted of an Agilent 1100 series high-performance liquid chromatography system (Agilent, Palo Alto, CA) with a Packard 525 Radiomatic flow scintillation analyzer (Packard Instrument Co.) and a Finnigan LCQ Deca XP⁺ ion trap mass spectrometer (Finnigan, San Jose, CA) as detectors. The chromatographic separation was achieved with an xTerra MS C₁₈ column, 250 by 4.6 mm, with a 5-µm particle size (Waters Corporation). The mobile phase A comprised 0.1% acetic acid containing 5% acetonitrile, and the mobile phase B was acetonitrile containing 0.1% acetic acid. The gradient conditions were 18 to 50% mobile phase B over 50 min and then to 100% over 20 min at 1 ml/min. The post column flow was split in a 1:20 ratio via an Acurate flow splitter (LC Packings, San Francisco, CA); 20 parts were sent to the flow scintillation analyzer and 1 part was sent to the ion trap mass spectrometer. The flow scintillation analyzer was equipped with a 250-µl flow cell, and the detection window was 4 to 100 keV. Packard Ultima FLO-M was used as the scintillation cocktail, and the flow rate was 2.5 ml/min. The LCQ mass spectrometer was equipped with an electrospray ion source and operated in positive mode. Key operating parameters included a spray voltage of 5 kV, a sheath gas flow of 35 U, an auxiliary gas flow of 0 U, a capillary temperature of 300°C, and an MS-MS collision energy of 40% with MS-MS wide excitation.

Pharmacokinetic analysis. The pharmacokinetic profiles of TPV were determined using plasma and blood TPV concentrations in a noncompartmental model (WinNonlin; Pharsight Corporation, Mountain View, CA). The following pharmacokinetic parameters for TPV in plasma and blood were analyzed: morning and evening trough concentrations, the C_max, the concentration in plasma 12 h after dosing (C_12h), the AUC, the time to maximum concentration (T_max), and the t_1/2 of total ¹⁴C radioactivity.

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Mass balance and excretion of radiolabel. A total of nine male subjects (eight white, one black) from 22 to 53 years of age with body mass indices of 21 to 27 kg/m² received radiolabeled TPV. Approximately 75.3% ± 23.7% (mean ± standard deviation [SD]) of the radiolabeled dose was recovered from feces, urine, and tissue (Table 1; Fig. 2, inset). Subject 113 was excluded from the mass balance analyses because diarrhea prevented complete sample collection. Subject 109 had 21.3% of the radiolabeled dose recovered, and this result was an outlier for the study group as a whole. For the remaining seven subjects, the total recovery of radioactivity was 83.0% ± 10.0%. Feces represented the primary route of excretion, with 70.4% ± 24.0% of the total radioactivity dose recovered via this route. Urinary excretion accounted for 4.6% ± 0.6% of the total administered radioactive dose.

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TABLE 1. Summary of cumulative recovery of radioactivity from samples from human subjects after oral administration of [14C]TPV

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FIG. 2. Radioactivity in plasma () and blood (•) after a single oral dose of [14C]TPV administered during the steady state. Means ± SDs are shown (n = 9). (Inset) Cumulative excretion of radioactivity in urine () and feces () and total radioactivity (•). Means ± SDs are shown (n = 8).

Radioactivity and TPV concentrations in plasma and blood. The pharmacokinetic parameters for TPV and for total radioactivity in plasma and blood are shown in Tables 2 and 3. Plots of mean concentrations of radioactivity in plasma and whole blood versus time are shown in Fig. 2. For all subjects, concentrations of radioactivity in plasma and blood declined steadily to below the limit of quantitation by 96 and 120 h postdose, respectively. The plots of blood and plasma radioactivity concentrations versus time were parallel, and the ratio of the concentration of circulating radioactivity in plasma to that in blood was 1.89 ± 0.08 at each time point. Therefore, approximately 85% of the TPV in blood was distributed in the plasma, with the remaining TPV in erythrocytes, and the distribution between plasma and blood cells was approximately constant. The plasma radioactivity-time profile could also be superposed on the plasma TPV concentrations obtained using the LC-MS-MS assay, indicating that the abundance of TPV relative to the total level of radioactivity in plasma remained approximately constant.

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TABLE 2. Pharmacokinetic parameters for TPV in plasma

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TABLE 3. Pharmacokinetic parameters for radioactivity in plasma and blooda

Pharmacokinetics. The pharmacokinetic profiles of TPV were characterized by using the concentration data for plasma as measured before and after the administration of the multiple-dose regimen and the single 551-mg oral dose of [¹⁴C]TPV. Pharmacokinetic data derived prior to the administration of radiolabeled TPV suggested that the steady-state plasma TPV concentration was achieved by day 7 of the twice-daily administration of TPV/r at 500 and 200 mg, respectively (Fig. 3). The range of trough plasma TPV concentrations for all subjects following the first two doses of TPV/r was 30.6 to 98.4 µM. TPV trough concentrations increased through three additional dose administrations with continued twice-daily dosing of TPV/r to a geometric mean of 70.0 µM on the evening of day 2, which was reflective of CYP3A inhibition by RTV. Mean evening trough concentrations decreased by 6 µM per day for the next 4 days, and by day 7 the trough TPV concentration was 21.9 µM. This level was indicative of the establishment of equilibrium between CYP3A inhibition by RTV and induction by TPV. A steady state was established by day 7, as the geometric mean ratio of the day 7 evening measurement to the day 8 morning measurement was 1.07. Geometric mean morning trough TPV concentrations from day 8 through day 15 averaged 18.2 µM (range, 16.6 to 21.9 µM).

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FIG. 3. Steady-state concentrations of TPV in plasma (Cp). Solid line, geometric mean; dashed line, median; day 1 to day 6, evening trough concentrations; day 7, morning and evening trough concentrations (evening concentrations were not normalized to the TPV dose of approximately 551 mg per subject); day 8 to day 15, morning trough concentrations (n = 7).

The calculated pharmacokinetic parameters for TPV are shown in Table 2, and those for total radioactivity are presented in Table 3. Using noncompartmental analyses, the median T_max was calculated as 3.0 h (range, 1.5 to 5.0 h), with a median normalized C_max of 83.56 µM (range, 67.87 to 128.31 µM) and a median normalized C_12h of 20.41 µM (range, 6.29 to 65.46 µM). The values were normalized for the actual radioactive dose (551 mg) given to the subjects as opposed to the nominal 500-mg dose given on the other days. The median elimination t_1/2 of TPV in plasma of 4.0 h (range, 2.8 to 7.3 h) closely approximated the median elimination t_1/2 of radioactivity in plasma of 3.96 h (range, 2.63 to 5.14 h) and that in whole blood of 4.19 h (range, 3.05 to 5.73 h). For this group of subjects, the median AUC from 0 to 12 h (AUC_0-12) following TPV/r administration at 500 and 200 mg was 545.5 h·µM (range, 409.9 to 840.6 h · µM).

TPV metabolites. Unchanged TPV was the predominant circulating component in the pooled plasma samples at 3, 8, and 12 h postdose and accounted for 98.4 to 99.7% of the plasma radioactivity (Table 4; Fig. 4a). Only a few trace metabolites, including a TPV glucuronidation conjugate (H-3) and an oxidation metabolite (H-1), were observed in radiochromatograms, and each metabolite accounted for 0.2% of the plasma radioactivity. Based on this observation of trace metabolites, it can be surmised that the plasma samples from all the subjects contained predominantly unchanged drug and that pooling was necessary for metabolite identification. Unchanged TPV also dominated the radiochromatograms for feces (Fig. 4c), accounting for 79.9% of fecal radioactivity or, on average, 53.5% of the dose in the nine subjects. Several metabolites were found in feces. Metabolite H-1, the most abundant, was responsible for 3.2% of the dose, while another oxidation metabolite, H-2, observed in the feces from most of the subjects, represented 0.4% of the dose. The remaining metabolites in feces each were less than 1.3% of the dose. In contrast, TPV was much less abundant in urine and represented only 0.5% of urine radioactivity or <0.1% of the dose (Fig. 4b). The majority of radioactivity in urine was accounted for by many metabolites. TPV glucuronide conjugate (H-3), accounting for 0.5% of the dose, was the most abundant urinary metabolite; the remaining urinary metabolites were each less than 0.4% of the dose.

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TABLE 4. Summary of abundance of radioactive metabolites in plasma, urine, and feces

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FIG. 4. Representative radiochromatograms for the 3-h plasma sample pooled from all nine subjects (a) and the urine (b) and feces (c) samples from subject 106 after the administration of the [14C]TPV dose.

The TPV glucuronide conjugate metabolite (H-3) corresponded to a radiochromatographic peak at approximately 52.5 min and a protonated molecular ion at m/z 779, 176 daltons higher than that corresponding to TPV. Upon collision activation, the molecular ion gave protonated TPV via the loss of glucuronide. The identification of H-3 was confirmed by a comparison of the retention time and the MS-MS pattern to the authentic standard.

Metabolite H-1 had a protonated molecular ion at m/z 619, 16 daltons higher than that of TPV (Fig. 5). MS-MS analysis of H-1 produced daughter ions at m/z 575 (loss of CO₂) and 411 via the same fragmentation patterns as TPV. In addition, new fragments were observed at m/z 557 (loss of CO₂ and loss of H₂O) and 495 (loss of C₇H₆O and loss of H₂O). The mass spectral analysis suggests that H-1 was likely a hydroxyl metabolite. Similarly, H-2 was tentatively identified as another oxidation metabolite, as illustrated in Fig. 5.

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FIG. 5. Comparison of MS-MS results for the TPV standard (a) and metabolites H-1 (b) and H-2 (c).

Safety. Nausea and loose stools were the most common adverse events associated with TPV/r administration. There were no serious adverse events, and only two subjects prematurely discontinued the use of the study drug, one due to nausea and alanine aminotransferase elevation and the other due to alanine aminotransferase elevation. The most common laboratory abnormalities were increases in transaminases (greater than or equal to Division of AIDS grade 3 in two subjects, or 17%) and mild increases in cholesterol and triglycerides.

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In this study conducted with healthy male volunteers dosed with twice-daily TPV/r at 500 and 200 mg, steady-state concentrations in plasma were achieved by day 7. Upon initial dosing, trough TPV concentrations were higher than those during the steady state, which is consistent with immediate CYP3A inhibition by RTV (5, 8). Trough TPV concentrations steadily decreased with twice-daily dosing until day 7 and thereafter remained relatively constant, reflecting steady-state attainment, through the remainder of the study (day 15). The progressive decrease in the trough TPV concentration and subsequent plateau suggest either CYP3A induction by TPV and RTV or a decreased bioavailability of TPV and RTV, followed by a balance of the discordant effects of TPV and RTV on metabolism and absorption. The trough TPV concentrations and other pharmacokinetic characteristics observed in this trial are consistent with those characterized in other clinical studies of TPV at similar TPV/r doses (8, 9).

After the administration of the [¹⁴C]TPV/r dose on day 7 (steady state), TPV was rapidly absorbed, and concentrations of both TPV and the radioactivity in plasma peaked at around 3 h postdose. TPV was found to be the predominant circulating component in plasma, accounting for more than 98.4% of plasma radioactivity at 3, 8, and 12 h postdose.

Approximately 75.3% of the radiolabeled dose was recovered from feces, urine, and tissue combined, and most was excreted by 96 h after dosing. In general, the level of recovery was lower than expected, perhaps as a result of incomplete sample collection by subjects. Fecal excretion, which accounted for approximately 70.4% of the radioactive dose, was the primary elimination route for TPV. On the other hand, urinary excretion was responsible for 4.6% of the radioactive dose and represented only a minor route of elimination. The absolute bioavailability of TPV, in the presence of RTV, is not known. The predominant recovery from feces in this study may indicate extensive biliary excretion or pronounced P glycoprotein efflux during the steady state. Although several metabolites existed in feces, each represented 3.2% of the dose. Additionally, all metabolites in urine were minor, with each accounting for 0.5% of the dose. The proposed metabolic pathways of TPV, in the presence of RTV, are shown in Fig. 6. The minor contribution of the kidneys to the elimination of TPV suggests that adjustments of the TPV/r dosage would be unlikely to be necessary for renally impaired patients.

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FIG. 6. Proposed metabolic pathways of TPV administered with RTV. Gluc, glucuronide.

The lack of significant TPV metabolism during the steady state in the presence of RTV coadministration and the significant decline in trough TPV concentrations from the initial dosing until the steady state was attained on day 7 suggest that the fractions of TPV and RTV absorbed are lower during the steady state than after a single dose. TPV (11) and, to a lesser extent, RTV (7) are substrates for P glycoprotein efflux transport. In a drug interaction study with the P glycoprotein substrate loperamide, TPV exhibited P glycoprotein self-induction capability (11). In long-term clinical trials with TPV and RTV, TPV has been shown to significantly reduce systemic RTV exposure (14), presumably due to P glycoprotein efflux transport.

This study demonstrated that the vast majority of the administered dose of TPV remains unchanged in plasma and excrements. The lack of TPV metabolism after the administration of TPV with RTV would be expected because of the profound inhibition of CYP3A by RTV (5). Previous studies have suggested that the first-pass metabolism of TPV, without RTV coadministration, occurs via the CYP3A pathway (8, 9), producing a maximum exposure of plasma to TPV of less than 1 µM. In addition, this study did not identify any major, unexpected TPV metabolites from an alternative pathway that might potentially have altered the pharmacokinetics, efficacy, and safety of TPV.

The major findings of this study, the unchanged nature of the majority of the administered dose in the plasma and the lack of any major, unexpected TPV metabolites, indicate that the structure of TPV remains intact and available to bind to HIV protease. The favorable TPV pharmacokinetic and metabolite profiles demonstrated in this study, along with the distinct structure and resistance profile of TPV, support TPV/r as an important treatment option for HIV-infected patients with resistance to multiple PIs.

 
* Corresponding author. Mailing address: Research and Development, Boehringer Ingelheim Pharmaceuticals, Inc., 900 Ridgebury Rd., Ridgefield, CT 06877. Phone: (203) 778-7870. Fax: (203) 791-6003. E-mail: lchen{at}rdg.boehringer-ingelheim.com

Published ahead of print on 7 May 2007.

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Antimicrobial Agents and Chemotherapy, July 2007, p. 2436-2444, Vol. 51, No. 7
0066-4804/07/$08.00+0     doi:10.1128/AAC.01115-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

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